Translocator protein (TSPO) is an 18 kD protein that is known to be mainly localised in peripheral tissues and glial cells but its physiological function remains to be clearly elucidated. Subcellularly, TSPO is known to localise on the outer mitochondrial membrane, indicating a potential role in the modulation of mitochondrial function and in the immune system. It has furthermore been postulated that TSPO is involved in cell proliferation, steroidogenesis, calcium flow and cellular respiration.
In studies examining the expression of TSPO in normal and diseased tissue, Cosenza-Nashat et al (2009 Neuropathol Appl Neurobiol; 35(3): 306-328) confirmed that TSPO expression in normal brain is minimal. This same paper demonstrated that in disease states elevated TSPO was present in parenchymal microglia, macrophages and some hypertrophic astrocytes, but the distribution of TSPO varied depending on the disease, disease stage and proximity to the lesion or relation to infection. Microglia and macrophages are the predominant cell type expressing TSPO in diseased brains, and astrocytes can also express TSPO in humans.
Ligands having affinity for TSPO are known in the art. A class of indole compounds having affinity for TSPO (IC50 values for most active compounds of between 0.2 nM and 5.0 nM) is disclosed in U.S. Pat. No. 6,451,795 as useful for the prevention or treatment of peripheral neuropathies and for the treatment of central neurodegenerative diseases. Okubu et al (Bioorg Med Chem 2004; 12: 3569-80) describe the design, synthesis and structure of a group of tetracyclic indole compounds having affinity for TSPO (IC50 values as low as about 0.4 nM).
A class of labelled tetracyclic indole derivatives was reported by Arstad et al (WO 2007/057705) as having nanomolar affinity for TSPO and therefore suitable for in vivo imaging of TSPO, e.g. in conditions such as Parkinson's disease, Alzheimer's disease, multiple sclerosis, neuropathic pain, arthritis, asthma, atherosclerosis and cancer. Radiolabelled tricyclic indole derivatives were disclosed by Wadsworth et al (WO 2010/109007) and reported to have nanomolar affinity in an in vitro assay as well as good metabolic stability and uptake in the brain in vivo sufficiently high and specific to indicate suitability of these compounds for application in in vivo imaging of TSPO expression in the central nervous system (CNS). Wadsworth et al (Bioorg Med Chem Letts 2012; 22: 1308-1313) and Achanath et al (WO 2011/117421) went on to report that these properties for in vivo imaging of TSPO in the brain were even more favourable in the S-enantiomer as compared to the racemate.
Purification of separated enantiomers of the above-described cyclic indole derivatives has presented challenges for the present inventors, in particular when trying to reproducibly obtain the purified enantiomer in a solid form when gram-sized batches are being processed. The present inventors have encountered difficulties when trying to obtain separated enantiomer in good quality solid form using the known rotary evaporation method, even when different solvents and/or different rotary evaporation equipment and/or rotary evaporation conditions are tried. There is therefore a need for an improved method for the purification of these compounds.